Patent GB-893880-A (GB)
Title: A new antibiotic a 9578 and pharmaceutical compositions containing same
Filed: 1958-07-28
Abstract: <PICT:0893880/IV (b)/1> A new antibiotic A9578 having components A and B, is prepared by culturing Streptomyces griseoflavus A9578, or a mutant thereof, under aerobic conditions in an aqueous nutrient medium containing a source of carbon, nitrogen and inorganic salts. The temperature is 18 DEG C.-40 DEG C. and preferably 27 DEG C. and the duration of fermentation is 1 1/2 to 5 days.  Typical sources of (a) carbon; are glucose, saccharose, lactose, mannitol, starch and glycerol, (b) nitrogen; are amino acids, peptides and proteins, peptones, tryptone, meat extract, cereal grain extract; distillation residues from alcohol, yeast, beans and cottonseed; ammonium salts and nitrates, and (c) inorganic salts are chlorides, carbonates, sulphates or the alkali metals, alkaline earth metals, magnesium, iron, zinc and manganese.  The fermentation broth is clarified at pH 3 to 5 and the mycelium washed with water or an aqueous organic solvent, e.g. aqueous methanol.  The antibiotic is isolated from the clarified broth by a combination of the following methods: (A) Adsorption on active carbon, activated earth or a resin such as a copolymer of m-phenylene-diamine and formaldehyde, and elution with aqueous acid, e.g. acetic or a mixture of water and a water miscible organic solvent, e.g. aqueous pyridine, or an aqueous mixture of such organic solvents.  (B) Adsorption on a cation exchange resin, e.g. "Amberlite" IRC 50 (Registered Trade Mark) in acid or sodium or mixed acid/sodium form, and elution with a dilute acid. (C) Precipitation as an insoluble salt with picric acid, picrolonic acid, 41-hydroxy-azobenzene-4-sulphonic acid or Reinecke acid.  Other salts are produced by treating the picrate with a salt of an organic base, e.g. triethyl-ammonium sulphate or with a dilute acid in an aqueous medium, e.g. aqueous methanol. (D) Addition of watermiscible solvent, acetone or dioxane to an aqueous or alcoholic aqueous solution of a salt of the antibiotic.  (E) Extracting an aqueous solution of the antibiotic with a solution of phenol in chloroform, the pH of the aqueous solution and the phenol content of the chloroform solution being varied, the antibiotic being recovered from the organic phase. (F) Adsorption chromatography on carbon, alumina, magnesium silicates, silica gel, calcium sulphate and ion exchange resins. (G) Distribution chromatography on cellulose, starch, silica gel and "Celite" (Registered Trade Mark) especially using a mixture of butanol/glacial acetic acid/water (4 : 1 : 5).  (H) Countercurrent distribution between two immiscible solvent phases, e,g. N/10 ammonium acetate buffer of pH 4.6 and 10% phenol in chloroform. A9578 base is obtained from the sulphate by reaction with barium hydroxide.  Salts are formed with inorganic and organic acids, e.g. hydrochloric, sulphuric, phosphoric, acetic, propionic, valeric, palmitic, oleic, succinic, citric, mandelic, pantothenic, glutamic and other amino acids; it is active against many microorganisms; has low toxicity; is stable in aqueous solution outside the range of 7 to 11.  Antibiotic A9578 is an iron and nitrogen containing basic compound of molecular weight from 1,000 to 1,050; readily soluble in water, methanol, dimethyl formamide and ethylene glycol monomethyl ether, sparingly soluble in ethyl acetate, ether, chloroform, dioxane, butanol and benzene; and having the following Rf values: 0.32 in butanol-pyridine-water (6 : 4 : 3) and 0.74 in butanol-ethanol-water (1 : 1 : 2).  Antibiotic A9578 has two components A and B having Rf values of 0.34 and 0.58, respectively, in water-saturated butanol containing 2% p-toluenesulphonic acid. Component A is an orange-yellow powder of empirical formula C42\sB1 H69\sB1 O15 N10 Fe, showing in the ultraviolet spectrum in aqueous solution maxima at <FORM:0893880/IV (b)/1> and an inflexion at <FORM:0893880/IV (b)/2> and exhibiting the infra-red spectrum shown in the Figure.ALSO:A pharmaceutical preparation for enteral, parenteral or local administration comprises antibiotic A9578, its component A, its component B, a salt thereof with an organic or inorganic acid, or a mixture thereof (see Group IV (b)) in admixture with an organic or inorganic carrier.  The preparations may be in the form of tablets, dragees, powders, salves, creams, suppositories, solutions, suspensions and emulsions.  They may contain preserving, stabilizing, wetting or emulsifying agents and other drugs, Suitable salts are the hydrochloride, sulphate, phosphate, acetate, propionate, valerate, palmitate, oleate, succinate, citrate, mandelate, pantothenate and glutamate.